Phylogenetic Evolution And Quantitative Genetic Analysis Of Major Traits For Chinese Vegetable Mustard (Brassica Juncea)

Mustard crop(Brassica juncea(L.) Czern.),belonging to the genus Brassica in the Cruciferae family,is an agriculturally and economically important crop widely cultivated in Asia and Europe.In India,B.juncea is used as oil-bearing crop and has been studied extensively.China is considered the original region of varietal differentiation,with the highest level of differentiation around Sichuan san Zhejiang.Chinese vegetable mustard has been increasingly attractive to biologist and plant breeders largely due to its higher diversification and economically important values.According to the U-triangle,B.rapa(2n=20,AA) and B. nigra(2n=16,BB) were hybridized naturally to synthesize the present species of B.juncea (2n=36,AABB).Although most studies on the species of U-triangle have confirmed the diploid origins of amphidiploid species,many questions remain concerning how the amphidiploid species evolved from their parental diploids.My research was focus on the phylogenetic analysis of B.juncea and genetic analysis of important characters on Chinese vegetable mustard.The main contents and results were as follows:(1) We applied genomic in situ hybridization(GISH) in this study that has allowed assignment of B.juncea chromosomes to the A or B genomes.The simultaneous application of B.nigra(red) and B.rapa(green) total genomic DNA probes allowed unambiguous discrimination of all 16 and 20 chromosomes of the genomes B and A,respectively.This demonstrates that despite their evolutionary proximity,a majority of repetitive DNA families from these two genomes do not share any significant level of homology.A detailed comparison of different A and B genomes revealed that the hybridization pattern of the B genome nuclear DNA differs from that of A genome.In the chromosomes of the AABB genome of B.juncea,although the GISH signals are localized predominantly in the pericentromeric regions,clear extensions of the signals towards the intercalary parts of the arm can be seen.In contrast,in the A genome,the signals are restricted almost exclusively to the pericentromeric regions;this may suggest that some differences in the distribution of repetitive DNA between the B genome and A genome.The results of most other cytogenetic (Harrison and Heslop-Harrison,1995) and molecular(Song et al.,1988;Warwick and Black, 1993) studies clearly suggest that these differences are not only quantitative but also qualitative.This result also supported the hypothesis that the B genome was the first to diverge from the common ancestral Brassica genome(Quiros,1995).(2) Sequence variation of nuclear internal transcribed spacer regions of ribosomal DNA (ITS1,5.8S rRNA and ITS2) from Chinese vegetable mustards(AB-genome) and its putative parents Brassica rapa(the A-genome) and Brassica nigra(the B-genome) were used to investigate the molecular phylogeny and the probable evolutional pattern of this amphidiploid species that uniquely formed in China.Totally,sixteen accessions of Chinese vegetable mustard those covering nearly all the diverse variations were included in this study, and together with three accessions of B.rapa and one accession of B.nigra.The results disclosed two strongly supported clades,one containing four accessions of vegetable mustard which have closer relationship with B-genome species "B.nigra" lineage and the other containing twelve accessions of B.juncea and three A-genome accessions.This classification was in disagreement with the evidence from chloroplast DNA,mitochondrial DNA,nuclear DNA restriction fragment length polymorphism(RFLP),which suggested that B.juncea was closely related to the A-genome type.For the incongruence,we speculated that the B.juncea crops derived from Chinese have evolved through different recombined events of the diploid morphutypes and evolved unidirectional concerted evolution.The traditional phenotypic classification of B.juncea was not wholly supported by ITS results,and hence the phylogenetic relationships among these subspecies need to be reconsidered on molecular level.(3) trnL intron and trnL-F were highly conserved among B.juncea species,however, nucleotide mutation points were found between B.rapa and B.nigra species.Informative sites in these two cpDNA regions povided evidence of the maternal origin of B.juncea.It was found that the materal parent of B.juncea could be B.rapa because the B.juncea were closely related to the accession of B.rapa (4) Amplified fragment length polymorphism(AFLP) analysis was used to evaluate the genetic relationships and diversities of Chinese vegetable mustards.The cluster analysis showed that the vegetable mustards could be grouped into two main groups and some minor rami,which was partially in accordance with the traditional classification that based on different edible organs of vegetable mustards.The incongruity between morphological and molecular classification might be attributed to the high selection pressure during domestication of Chinese vegetable mustards,producing some accessions with similar genetic backgrounds evolving into abundant morphological variations.The great diversification among Chinese vegetable mustards not only provides an excellent object for molecular evolution research of B.juncea but also is of great value for widening the genetic basis of breeding programs and breeding materials selection.Besides,our study also indicates that AFLP are informative and can provide significant insights for genetic diversity research in B.juncea.(5) Genetic effects and genotype by environment(GE) interaction effects for some important agronomic traits and qualitative traits of Chinese vegetable mustard were analyzed by using a genetic model including additive,dominance,additive×additive effects and their interaction effects with environments.Four different variations of Chinese vegetable mustard as parental lines and their F_1s,F_2s were evaluated in fields of two locations.It was revealed that the agronomic traits of Chinese vegetable mustard were mainly controlled by genetic effects except plant weight(PW) and leaf weight(LW) were observed to be more affected by GE interaction effects.Among the genetic effects,additive effects took the main proportion for tillering number(TN),leaf number(LN),leaf breadth (LB) and LW;dominance effects were the main components of PW,leaf length(LL),root weight(RW) and plant height(PH);additive×additive effects were the main components of plant breadth(PB).Among the GE interaction effects,LB,LW and RW were mainly affected by additive interaction effects and PW,LL,PH and PB were mainly controlled by dominance interaction effects.Besides,additive×additive interaction was the main factor which controlled TN and LN of Chinese vegetable mustard.For heterosis analyses,TN,LN, LB and LW of Chinese vegetable mustard showed positive H_PM and negative H_PB.The other traits showed positive H_PM and H_PB.Heterosis arising from GE interaction was found to varying degree for different locations.It was shown that genetic heterosis and GE interaction effects were important factors for agronomic traits in Chinese vegetable mustard.For quality traits,it was revealed that Vitamin C content and total protein content were mainly controlled by additive effects and additive environment interaction effects.Sugar content,amino acid content,sinigrin content and total glucosinolate content were mainly controlled by dominance and dominance environment interaction effects.For heterosis analyses,sinigrin content and total glucosinolate content of Chinese vegetable mustard showed positive H_PM and negative H_PB The sugar content and total protein content showed negative H_PM and H_PB.Vitamin C content showed negative H_PB although no H_PM.Amino acid content of Chinese vegetable mustard showed positive H_PM(F₁) and negative H_PM(F₂) and H_PB.